Aryl Hydrocarbon Hydroxylase Research Articles

Microsomal and cytosolic cytochrome P450 mediated benzo(a)pyrene hydroxylation in Pleurotus pulmonarius

Microsomal and soluble fractions of Pleurotus pulmonarius exhibited a reduced carbon monoxide difference spectrum with P450 maxima at 448nm and 450–452nm respectively. Substrate induced Type I spectra were observed on addition of benzo(a)pyrene to both fractions. Benzo(a)pyrene hydroxylation was measured using the aryl hydrocarbon hydroxylase assay and was observed to be P450 dependent as indicated by carbon monoxide inhibition together with the substrate binding characteristics. The activity of the fractions were observed to give Km of 200mM and 660mM and Vmax of 1.25 nmol/min/nmol P450 and 0.57 nmol/min/nmol P450 for the microsomal and cytosolic fractions respectively.

Influence of growth hormone on the hepatic mixed function oxidase and transferase systems of rainbow trout

The effect of GH treatment on hepatic cytochrome P450 content, aryl hydrocarbon hydroxylase (AHH), aminopyrine-N-demethylase (AND), testosterone hydroxylase, testosterone 5α- and 5β-reductase, UDP-glucuronyl transferase (UDPGT) and glutathione S-transferase (GST) activities in immature rainbow trout were investigated. Hepatic cytochrome P450 content, AHH and GST activities were measured in both GH implanted and GH injected animals whereas other activities were assayed in GH implanted trout only.GH implants significantly decreased cytochrome P450 content at 15 days compared to the control but no significant effect was observed at 15 or 30 d when GH was injected biweekly. In both cases, AHH activity was significantly decreased by GH treatment compared to the control whereas GST remained unchanged. Compared to the control, GH implanted fish exhibited a pronounced inhibition of AND, a decreased 6β and 16β-testosterone hydroxylation, an inhibition of UDPGT with testosterone as substrate and an enhanced 17β-testosterone oxidation.

Different Inactivation Mechanisms of NADPH- and NADH-Dependent Chains of Rat Liver Endoplasmic Reticulum in Tetrachloromethane

Cobalt-protoporphyrin (CoPP) administration to adult male rats results in a profound reduction in hepatic cytochrome P-450 concentration and activity, and decreased plasma concentrations of testosterone and luteinizing hormone (LH). The metabolism of progesterone by rat testicular microsomes isolated 48 h after treatment in vivo with CoPP was compared to that in microsomes from control rats. The conversion of progesterone to 17α-hydroxyprogesterone and 4-androstenedione, which is NADPH-dependent, was reduced by approximately 40% in testicular microsomes following treatment with CoPP (50 μmol/kg body weight) and this inhibition was dose-dependent. The concentration of cytochrome P-450 in testicular microsomes and the activity of 7-ethoxycoumarin de-ethylase (a cytochrome P-450 dependent function) were also reduced following treatment with CoPP in contrast to two other functional assays of cytochrome P-450, aryl hydrocarbon hydroxylase and ethylmorphine demethylase, which were unaffected by treatment with CoPP. Thus, the profound effect of CoPP on androgen homeostasis has been extended to include decreased testicular synthesis of 4-androstenedione in addition to increased hepatic metabolism of testosterone, attenuated pituitary LH release in response to luteinizing hormone-releasing hormone, and failure of testicular response to LH.

Biochemical and Biophysical Characterization of Pulmonary Surfactant in Rats Exposed Chronically to Cigarette Smoke

Biochemical and Biophysical Characterization of Pulmonary Surfactant in Rats Exposed Chronically to Cigarette Smoke. Subramaniam, S., Bummer, P., and Gairola, C. G. (1995). Fundam. Appl. Toxicol. 27, 63-69. The pulmonary surfactant plays an important role in the gas exchange functions of the lungs. Although previous studies suggest that cigarette smoking alters the pulmonary surfactant system in human smokers, the nature of such changes is poorly understood. The aim of the present study was to determine if biochemical and biophysical properties of pulmonary surfactant are affected in rats following chronic exposure to cigarette smoke. Female Sprague-Dawley rats were exposed daily to smoke from the University of Kentucky high tar/high nicotine reference cigarettes, twice a day, for 60 weeks in a nose-only exposure system. Blood carboxyhemoglobin, plasma cotinine, and pulmonary aryl hydrocarbon hydroxylase activity measurements showed that animals effectively inhaled smoke during exposures. At termination, the bronchoalveolar lavage fluids (BALF) and the lung tissues were collected for biochemical and biophysical analyses of surfactant. The total phospholipid content of the BALF and the lung tissues from room control (RC), sham-treated (SH), and smoke-exposed (SM) animals were the same among the different groups. However, disaturated phosphatidylcholine (DSPC) levels in the BALF were significantly decreased in SM rats compared to RC or SH groups. In contrast, the lung tissue DSPC content in SM rats was not significantly different from that of control groups. Phospholipid profile analysis of the BALF also did not reveal any significant differences among other major constituents of surfactant from control and SM animals. The organic extracts of BALF obtained from different animal groups were assessed for surface activity using a Wilhelmy balance. The results showed an increase in surface compressibility and a reduction in respreadability index in SM group. In contrast, the stability indices of surfactant films obtained from different groups were the same. These observations suggest that decreased DSPC content in SM rats plays a role in altering the biophysical characteristics of surfactant. It is concluded that chronic exposure to cigarette smoke adversely affects the pulmonary surfactant system of rats.

In vivo murine studies on the biochemical mechanism of acetaminophen cataractogenicity

C57BL/6 and DBA/2 mice are, respectively, susceptible and resistant both to the induction of aryl hydrocarbon hydroxylase (cytochrome P450 1A1, or CYP1A1) and to the cataractogenicity of acetaminophen, which may involve its bioactivation to a toxic reactive intermediate, catalysed by P450 and (or) prostaglandin H synthase (PHS). Following induction of P450 using beta-naphthoflavone, the cataractogenicity of acetaminophen (400 mg/kg ip) in C57BL/6 mice was reduced by pretreatment with the P450 inhibitors SKF 525A and metyrapone, the glutathione precursor N-acetylcysteine, the antioxidant vitamin E, and the free radical spin trapping agent alpha-phenyl-N-t-butylnitrone (p < 0.05). Acetaminophen (200 mg/kg) cataractogenicity was enhanced by pretreatment with the glutathione depletor diethyl maleate (DEM) and the gamma-glutamylcysteine synthetase inhibitor buthionine sulfoximine (BSO) (p < 0.05). No significant effect on acetaminophen cataractogenicity was observed using the PHS cyclooxygenase inhibitors aspirin or naproxen, or the glutathione reductase inhibitor 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). Accordingly, acetaminophen cataractogenicity in C57BL/6 mice does not appear to be dependent upon bioactivation by PHS. In DBA/2 mice treated with beta-naphthoflavone, a high dose of acetaminophen (750 mg/kg ip) was not cataractogenic, even after pretreatment with DEM, BSO, or BCNU. The resistance of DBA/2 mice to acetaminophen cataractogenesis, despite concomitant pretreatments with an inducer of P450 and several agents that interfere with glutathione-dependent detoxifying pathways, suggests differences in this strain involving cytoprotective pathways subsequent to acetaminophen bioactivation and detoxification of the cataractogenic reactive intermediate. These results indicate that acetaminophen cataractogenicity in C57BL/6 mice results from P450-catalysed bioactivation of acetaminophen to a reactive intermediate, possibly a benzoquinone imine and (or) a free radical, the toxicity of which is reduced by glutathione-dependent reactions.

Nitroreduction of 2,4-dinitrotoluene [formula omitted] by cytochrome P-450 induced H4IIE cells

Conditions have been established for H4IIE rat hepatoma cell cultures in which effects of cytochrome P-450 induction on the metabolism of a munitions wastestream pollutant can be studied. Under these conditions, the polychlorinated hydrocarbon 2,3,4,7,8-pentachlorodibenzfuran (PCDBF) induced cytochrome P-450 (1A1) aryl hydrocarbon hydroxylase (AHH) activity over a wide range of concentrations without significant cytotoxic effects. The munition pollutant 2,4-dinitrotoluene (2,4-DNT) did not induce AHH activity itself, but its metabolism was considerably altered when applied to PCDBF induced cultures. Production of amino nitrotoluene isomers was greatly enhanced in induced cultures as compared to uninduced controls, as was the conversion of radiolabeled 2,4-DNT to relatively more polar metabolites. To some extent, the results with H4IIE cells parallel those reported for animals exposed to 2,4-DNT after induction of cytochrome P-450 AHH activity. The preliminary findings suggest that with further development and validation, H4IIE cultures could be of use in characterizing metabolites that result from exposure to chemical mixtures involving a P-450 (1A1) inducer.

Polymorphisms of human Ah receptor gene are not involved in lung cancer.

The Ah receptor (Ahr) is a ligand-dependent transcription factor that positively regulates inducible expression of the CYP1A1 gene. Based on the sequence information of the human Ahr and the intron-exon junctions of the mouse counterpart, an analysis of single-strand conformational polymorphism (SSCP) was carried out to detect subtle base differences in the coding region of the gene among individuals. We found that the Ahr protein has at least two forms of variants in a Japanese gene pool, and that these variants can be ascribed to one amino acid replacement of Arg by Lys at codon 554. The frequencies of Arg-coded and Lys-coded alleles were 0.57 and 0.43, respectively. We found, however, that this germ line polymorphism of the Ahr gene did not show a significant association with aryl hydrocarbon hydroxylase (AHH) inducibility nor with lung cancer incidence.

Metabolic activation and toxicity of some chemical agents to lung tissue and cells

Selected topics: Pulmonary bronchiolar epithelial cytotoxicity: microanatomical considerations, C. G. Plopper. Xenobiotic metabolism by isolated pulmonary bronchiolar and alveolar cells, T. R. Devereux et al . Localization, distribution, and induction of xenobiotic-metabolizing enzymes and aryl hydrocarbon hydroxylase activity within lung, J. Baron & J. M. Voi t. Purification and characterization of lung enzymes involved in xenobiotic metabolism, F. P. Guengerich. Action by the lungs on circulating xenobiotic agents, with a case study of physiologically based pharmacokinetic modeling of benzo(a)pyrene disposition, R. A. Roth & A. Vinega . Metabolism of endogenous and xenobiotic substances by pulmonary vascular endothelial cells, U. S. Ryan & A. P. Li. Naphthalene and 2-methylnaphthalene-induced pulmonary bronchiolar epithelial cell necrosis: metabolism and relationship to toxicity, R. B. Franklin et al . Pulmonary toxicity of 4-ipomeanol, T. E. Gram. Pulmonary toxicity induced by phosphorothioate impurities present in organophosphate insecticides, J. Gandy et al . The metabolic basis of 3-methylindole-induced pneumotoxicity, T. M. Bray & J. B. Kir land. Metabolism and pulmonary toxicity of butylated hydroxytoluene, H. Witschi et al . Hepatic nonaltruism and pulmonary toxicity of pyrrolizidine alkaloids, R. J. Huxtable. Cyclophosphamide: pulmonary metabolism, toxicity and protective effect of vitamin E, J. M. Patel. Metabolic activation and biological effects of nitrosamines in the mammalian lung, H. M. Schuller et al . The use of bleomycin in model systems to study the pathogenesis of interstitial pulmonary fibrosis, J. S. Lazo et al . The pulmonary toxicity of nitrosoureas, A. C. Smith. Index.

Hepatic and biliary biomarkers in rainbow trout injected with sediment extracts from the River Po (Italy)

Immature rainbow trout ( Oncorhyncus mykiss) were injected i.p. with varying doses of extracts of sediments collected from the River Po (Northern Italy), upstream and downstream the immission of a heavily contaminated tributary (River Lambro). Six days after treatment, metabolic biomarkers were monitored in fish liver and bile. Microsomal enzyme activities, including arylhydrocarbon hydroxylase, ethoxyresorufin O-deethylase, aminopyrine N-demethylase and uridine-5′-diphosphoglucuronyltransferase, were induced by exposure to the polluted sediment to a moderate yet statistically significant extent, with dose-related effects. The cytosolic enzymes glutathione reductase, glutathione peroxidase and glutathione S-transferase were not affected by injection of sediment extracts, whereas both upstream and downstream specimens produced a depletion of nonprotein sulfhydryl groups. Elimination of fluorescent metabolites occured in the bile of trout injected with the polluted sediment extract, whereas organic extracts of bile were devoid of mutagenic activity in strain TA98 of S. typhimurium. In spite of some positive responses, the method used appears to be less sensitive in revealing the toxicological impact of pollution than either exposure of rainbow trout larvae to river sediment or in situ exposure of the same fish species in river water.

Dioxin-Type Activity for Polyhalogenated Arylic Derivatives. A QSAR Model Based on MTD-Method

Abstract QSARs by the Minimal Steric/Topologic Difference (MTD)-method were performed for general toxicity, hepatic cytosol receptor binding and aryl hydrocarbon hydroxylase induction on polychlorinated and polybrominated derivatives of dibenzo-p-dioxin, dibenzofuran and biphenyl. Low energy conformations selected by molecular mechanics calculations and superpositions of the considered structures for the construction of the hypermolecule were performed using the COSMIC molecular modelling package. The multiconformational-MTD-method was used. Correlation coefficients of r = 0.8 to 0.9 were obtained as well as good cross-validation results, for series of up to 70 compounds. Conformational analysis of flexible molecules suggests their accommodation in the receptor site also in non planar conformations corresponding to the first stable one (minimum energy costs) above the global minimum. A common optimised receptor site can be attributed for all series of compounds, with a rather high symmetry, which suggests...

Differential effects of dietary diallyl sulfide and diallyl disulfide on rat intestinal and hepatic drug‐metabolizing enzymes

The chemopreventive properties of allyl sulfides on carcinogenesis may be related to the modulation of drug-metabolizing enzymes involved in carcinogen activation or detoxication. In order to investigate the effects of diallyl sulfide (DAS) and diallyl disulfide (DADS) on intestinal and hepatic drug-metabolizing enzymes, rats were fed a diet containing 0.2% of either allyl sulfide. The DADS enhanced intestinal epoxide hydrolase (EH) and cytochrome P-450 (P-450) 2B1/2 protein levels and the activities of pentoxy- and benzyl-oxyresorufin O-dealkylases, arylhydrocarbon hydroxylase, microsomal epoxide hydrolase, p-nitrophenol UDP-glucuronyl transferase and glutathione S-transferase, and decreased nitrosodimethylamine demethylase activity. In liver, DADS produced similar effects and, in addition, increased P-450 1A1/2 protein level and phenoxazone metabolizing activities (ethoxy- and methoxyresorufin O-dealkylases), p-hydroxybiphenyl UDP-glucuronyl transferase, and decreased P-450 2E1 level. The DAS enhanced only EH activity in the small intestine and induced P-450 2B1/2 and epoxide hydrolase protein levels. In liver, DAS produced similar effects as DADS. The different effects of DAS on intestinal drug-metabolizing enzymes, compared to liver, could be ascribed to less metabolism of this compound in small intestine. It is also suggested that DAS and DADS may not yield the same metabolites and therefore would have different effects on intestinal drug-metabolizing enzymes.

Comparison of expression of aldehyde dehydrogenase 3 and CYP1A1 in dominant and recessive aryl hydrocarbon hydroxylase-deficient mutant mouse hepatoma cells

The mouse hepatoma cell line Hepa-1 is inducible by 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) for both CYP1A1 (aryl hydrocarbon hydroxylase, AHH) and class 3 aldehyde dehydrogenase (ALDH3) enzymes. To test the hypothesis of a common regulatory mechanism, several AHH deficient mutants of Hepa-1 were studied for their ALDH3 activities and specific mRNA levels before and after TCDD treatment. The recessive (with respect to the wild-type Hepa-1) mutants have defects in Cypla-1 structural gene (mutant c1) or in the Ah (aryl hydrocarbon) receptor (mutants c2 and c6 with decreased levels of Ah receptor; mutant c4 defective in the DNA binding of the Ah receptor). The results with these mutants suggested that Ah receptor nuclear translocator protein, ARNT, is needed for ALDH3 expression. Two dominant mutants, one of which is characterized by preventing the binding of the Ah receptor complex to DNA, were also studied. Surprisingly, these mutants possessed elevated levels of ALDH3 mRNA and enzyme activities which were also inducible by TCDD. The binding of Ah receptor-ligand complex to DNA was thus not needed for the expression of ALDH3. A dominant repressor for Cypla-1 gene transcription did not prevent the derepression or induction of ALDH3. The results thus suggest that Aldh-3 gene is regulated by a mechanism independent of the Ah receptor.

Immunocytochemical study of cytochrome P450 (1A1/1A2) induction in murine ocular tissues

C57BL/6 and DBA/2 mice were injected intraperitoneally with beta-naphthoflavone in corn oil and killed 48 hr later. Control animals received an injection of corn oil. The immunoreactivity of cytochrome P450 1A1/1A2 expressed in different ocular tissues and liver was examined with goat anti-P450 antibody (primary antibody) and gold-conjugated anti-goat antibody (secondary antibody). DBA/2 mice, which are non-responsive to aryl hydrocarbon treatment, showed negligible levels of immunoreactivity toward anti-P450 1A1/1A2 antibody in all ocular tissues, whether or not the animals were treated with beta-naphthoflavone. In responsive C57BL/6 mice, however, the immunoreactivity of the uveal tissues, especially ciliary non-pigmented epithelium, was markedly increased by beta-naphthoflavone treatment. The time course of induction of P450 1A1/1A2 immunoreactivity was very similar for the liver and ciliary non-pigmented epithelium, although the maximum level of immunoreactivity of the ciliary epithelium reached in 48 hr after inducer treatment was about 25% of that of liver. The present results support our previous observations that the P4501A enzyme activities (e.g. aryl hydrocarbon hydroxylase) in the liver and eye of C57BL/6 mice are under the same genetic regulation. Further, this study is the first demonstration of P450 isoform induction in specific ocular tissues of the whole animal.

The relationship between age and cutaneous aryl hydrocarbon hydroxylase (AHH) activity.

Aryl hydrocarbon hydroxylase (AHH), a component of the mixed-function oxygenase system, has been identified in liver and in many extrahepatic tissues, including the skin. In this study, the effect of ageing on cutaneous AHH activity in both male and female Wistar and Brown Norwegian rats of two age groups (average ages being 44 and 130 weeks) was assessed using benzo(alpha)pyrene as substrate and by measuring the formation of the fluorescent metabolite 3-hydroxybenzo(alpha)pyrene. Results show that cutaneous metabolism of polycyclic aromatic hydrocarbons does not change between young adulthood and senescence in either male or female rats. Sex-related differences did occur, however, and may be explained, partly, by the existence of different molecular forms of cyt-P450. In conclusion, age does not appear to be a major determinant of basal cutaneous AHH activity in rats.

Aromatic hydrocarbon receptor in cultured fetal cells from C57BL/6J and DBA/2J mice: similarity in molecular mass to receptors in adult livers.

In liver of adult responsive C57BL/6J (B6) mice the aromatic hydrocarbon receptor (AHR) has high affinity for specific halogenated aromatic hydrocarbons, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), as well as nonhalogenated aromatic hydrocarbons (PAHs), such as benz[a]anthracene (BA) or 3-methylcholanthrene (MC). In livers of adult nonresponsive DBA/2J (D2) mice TCDD binds to a low-affinity variant form of AHR. Both TCDD and MC induce aryl hydrocarbon hydroxylase (AHH) in adult B6 mice, whereas adult D2 mouse liver is nonresponsive to MC. In fetal cell cultures derived from D2 mice AHH is induced by PAHs such as MC or BA, and these PAHs bind to cytosolic AHR (P.A. Harper, C.L. Golas, and A.B. Okey. Mol. Pharmacol. 40: 818-826, 1991). We compared AHR from fetal cell cultures with AHR from adult livers to determine whether there was some structural differences in receptors expressed in fetal cell culture that might permit cells from "nonresponsive" mice to respond to PAHs. The apparent molecular mass of AHR from cells cultured from 18-day fetuses is identical with that from adult liver within each strain of inbred mice tested (M(r) approximately 95 kDa in B6 and approximately 105 kDa in D2 mice). The AHR in D2 fetal cells was able to activate a transfected chloramphenicol acetyltransferase linked to a dioxin-responsive element nucleotide sequence (DRE-CAT) when the cells were treated with TCDD or MC. The potency of CAT expression in D2 fetal cells was similar to that in B6 fetal cells. Our data suggest that the responsiveness of fetal cells from "nonresponsive" mice is likely mediated by AHR in these cells but is not due to expression of a different allelic form of AHR ligand-binding subunit in fetal cells versus adult liver.

Expression and polymorphism of glutathione S-transferase in human lungs: risk factors in smoking-related lung cancer.

The relationships between smoking and the expression of glutathione S-transferase (GST*) isozymes GSTM1-1, GSTM3-3, GSTP1-1 and GSTA1-1/2-2 (GSTA1/2), or between smoking and activities of epoxide hydrolase (EH) and aryl hydrocarbon hydroxylase (AHH) were investigated in lung samples from 27 patients with lung cancer and 11 control patients by immunoblot analysis and enzyme assays. Determination of genotypes in blood leucocyte DNA showed that possession of the mu-class GSTM1 gene was closely related to the expression of GSTM1-1 and GSTM3-3 enzymes in lung cytosol: patients with the GSTM1 null genotype had no detectable GSTM1 protein and less GSTM3 protein than patients with the GSTM1 gene (P < 0.001). Absence of the GSTM1 gene did not affect the content of phi-class GSTP1-1 or alpha-class GSTA1/2. GST activity towards 1-chloro-2,4-dinitrobenzene was lower (P < 0.01) in patients lacking the GSTM1 gene than in those expressing GSTM1; in general, patients with a low GSTM3-3, GSTP1-1 or GSTA1/2 content also had significantly less overall GST activity. The pulmonary content of GSTP1-1 was greater in cancer than in non-cancer patients (P < 0.05). Smoking did not influence the levels of GST isozymes or the EH activity. In contrast, the AHH activity was significantly (P < 0.01) increased by smoking. Neither AHH nor EH showed a correlation with GSTM1 polymorphism. Our data support the idea that in smokers who lack the GSTM1 gene, activation of carcinogens in tobacco smoke (e.g. benzo[alpha]pyrene) is increased, while the efficacy of detoxification is limited both qualitatively (absence of GSTM1-1 enzyme and low expression of GSTM3-3 enzyme) and quantitatively (low overall GST activity). This imbalance in the metabolism of carcinogens may explain the increased susceptibility to lung cancer reported in smokers with the GSTM1 null genotype.

A field evaluation of cytochrome P4501A as a biomarker of contaminant exposure in three species of flatfish

AbstractA study was conducted over the course of a year to determine the induction of hepatic cytochrome P4501A (CYP1A) in three species of benthic fish collected from a contaminated site compared to fish sampled from a less‐contaminated site Juvenile fish were used to minimize effects of reproductive status and migration CYP1A was determined by two catalytic assays [aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin O deethylase (EROD)] and by an immunoassay (ELISA) utilizing polyclonal antibodies raised against purified CYP1A from cod AHH activities were measured by a standard method (AHHstd) and by two variations of the standard method All three primary CYP1A measures (AHHstd, EROD, and ELISA) showed consistent between site differences, indicating that induction of CYP1A can be a reliable biomarker of contaminant exposure in fish if appropriate biological variables are controlled for in field studies Multiple ANOVA demonstrated that the AHHstd and ELISA data showed less variability due to species or temporal differences, and less unexplained variability, compared to the data from the EROD assay or either variation of the AHH assay For all measures, variability associated with site differences far outweighed species or temporal variability Immunoassay, while less sensitive than the AHHstd assay, is nonetheless recommended to be used in conjunction with catalytic assays because of the potential for samples to lose catalytic activity if not handled properly The current results suggest that the lower noncontaminant related variability of AHHstd makes this CYP1A measure potentially more useful for monitoring programs in which analysis of trends is a primary goal

Induction of rat hepatic P4501A1 by organic extracts from airborne particulate matter in Santiago, Chile

1. Organic extracts from particulate matter collected in downtown Santiago, Chile, in 1990 were administered to female Wistar rats. 2. Extracts shifted the maximal absorption wavelength P450 spectra of the reduced-CO complex of hepatic microsomal P450 from 450 to 448 nm, and enhanced the total content of P450. In addition, substantial increases in aryl hydrocarbon hydroxylase, ethoxyresorufin O-deethylase and ethoxycoumarin O-deethylase activities were observed, whereas aminopyrine N-demethylase activity was not affected by treatment. 3. Western blotting using polyclonal antibodies against P4501A isozymes showed the appearance of a distinct and intense P4501A1 band in microsomes from rat pretreated with air particle extracts, and was not observed in microsomes from control rat. On the other hand, the intensity of the P4501A2 isoenzyme was apparently not affected. 4. These findings suggest that the organic extracts from airborne particulate matter modify the composition of rat liver P450 isozymes by inducing those isoforms responsible for the activation of precarcinogenic to carcinogenic agents.

Growth hormone effect on xenobiotic-metabolizing activities of rainbow trout

In recent years several studies reported the regulation by growth hormone (GH) of the expression of a variety of P450 forms in mammals. However the effect of GH on the xenobiotic-metabolizing enzymes of fish are still unknown. The aim of this work was to investigate the effects of ovine GH—a growth hormone known to be efficient in trout—on the cytochrome P450 level and on aryl hydrocarbon hydroxylase, aminopyrine-N-demethylase, glucuronyl transferase and glutathione transferase activities in trout. The GH-implanted trout (n = 50) each received a single cholesterol pellet containing ovine GH and were compared to control animals (n = 50) receiving a single cholesterol pellet without GH. After 15 days fish were killed and the liver and gills were excised for the measurement of xenobiotic-metabolizing enzyme activities. GH treatment significantly decreased the level of hepatic cytochrome P450 and the activities of cytochrome P450 dependent monooxygenases. In contrast, no significant effect of the treatment was observed on the glutathione transferase and UDP-glucuronyl transferase activities. Moreover, GH treatment had no effect on branchial phase I and phase II enzyme activities. This study provides evidence that GH level significantly affects the expression of several members of the hepatic cytochrome P450 family in trout.

Relationships between induction of cytochrome P4501A and contaminant residues in feral teleosts

We are currently analyzing hepatic cytochrome P4501A and associated monooxygenase activities in fish sampled in several regional and national monitoring programs, including the National Benthic Surveillance Project of NOAA's Status and Trends Program, damage assessment studies of the Exxon Valdez oil spill, and intensive surveys of specific embayments, such as Puget Sound, Washington. Thus far, apparent contaminant-related increases in the activities of cytochrome P4501A-dependent monooxygenases have been readily measured in most test species. The results presented in this paper show that, for II species of fish, there is excellent concordance between hepatic activities of aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin-O-deethylase (EROD); moreover, levels of cytochrome P4501A determined by an enzyme-linked immunosorbent assay (ELISA) are also generally concordant with results from catalytic assays. The use of both a catalytic assay and immunoquantitation is recommended, because of the additional quality assurance provided by concurrent use of an immunoquantitation technique, which is desirable in large monitoring programs.